Behavioural Processes

The vast majority of birds are diurnal and concentrate their vocal activity during daylight hours. However, some diurnal birds can also be vocally active at night, although the functions of this phenomenon remain poorly understood. We conducted playback experiments in the Warta Landscape Park (central Poland) to determine whether nocturnal singing by diurnal birds serves breeding-related functions by analysing responses to playback of songs from unfamiliar conspecific males. The sedge warbler (Acrocephalus schoenobaenus) was selected as the focal species because it exhibits relatively high levels of nocturnal song activity, while nine additional diurnal species detected near focal sedge warbler territories were included to explore whether responsiveness to nocturnal conspecific song extended across a broader taxonomic range. Playback experiments were conducted during the early and late stages of the breeding season and during the early and late parts of the nautical night. Out of 10 species tested, three responded vocally: sedge warbler, Savi s warbler (Locustella luscinioides), and common snipe (Gallinago gallinago). Sedge warblers did not modify song rate and song duration but increased flight activity after nocturnal playback. Savi s warblers and common snipes produced more vocalisations after playback than before in May, a pattern consistent with territorial defence function. General nocturnal vocal activity was higher at the beginning of the season, suggesting that birds motivation to establish territories and form pairs can extend into the night, providing additional benefits. Moreover, the probability of singing by the sedge warbler was higher in the latter part of the night. Our study demonstrates that nocturnal stimulation of foreign male playback of diurnal birds can elicit vocal responses from conspecifics, suggesting that nocturnal singing can occur in the absence of obvious artificial light pollution, but in the case of some species and environmental conditions, it may contribute to nocturnal social communication, especially in the early breeding season.

The ability of prey to eavesdrop on predator vocalizations is expected to increase survival by reducing detection and capture. Unfortunately, most research has been conducted in vertebrates, and little is known about this ability in invertebrates. We measured latency to emerge, overall activity, and shelter visits in wild-caught fall field crickets (Gryllus pennsylvanicus) in response to acoustic playback. Stimuli included multiple predator vocalizations, non-predator vocalizations, white noise, and a control. We predicted that crickets would reduce activity, spend more time in shelter, and freeze in response to stimuli representing greater risk. Contrary to our predictions, crickets traveled greater distances, spent more time moving, and spent less time in shelter in response to predator vocalizations versus controls. We did not, however, find clear differences in responses between predator vocalizations and other treatments. Our results suggest that crickets may not differentiate between the vocalizations of predators, non-predators, and other abrupt sounds. Consequently, eavesdropping may not be a viable method of assessing predation risk for this species and its general use remains unclear.

Animals experience stressors in a variety of contexts that result in activation of neuroendocrine and cellular stress responses. Release of stress hormones can disrupt or restore redox homeostasis, and the resulting changes in oxidative states, physiology and behavior vary by an individuals stress coping style. However, oxidative stress can also directly modulate neuroendocrine stress signaling. To what extent individual differences in brain antioxidant levels alter behavioral stress levels is not well understood. The present study investigated how N-acetylcysteine amide (NACA), an antioxidant and glutamate-modulating compound, regulates stress behavior across zebrafish (Danio rerio) with different stress coping styles (proactive, reactive). Following 24-hour exposure to NACA or control conditions, we quantified individual and composite stress behaviors using a Light-Dark Test (LDT). As expected, both proactive fish and NACA-treated fish showed significantly lower stress behaviors compared to reactive and control animals, respectively. Notably, stress-reducing effects of NACA were only seen in those with a reactive stress coping style. Overall, our data suggest that antioxidant mechanisms (e.g., glutathione system) may be key in facilitating the distinct behavioral and physiological responses to stressors that characterize alternative stress coping styles. The results underscore how individual differences in stress coping style and redox state can influence behavioral responses to stress.

Healthy lifestyle habits are a key pillar of physical, psychological, and social well-being, yet university years are often marked by declines in sleep quality, physical activity, and other health behaviors that may affect academic performance. This cross-sectional study examined these associations in 390 Spanish university students, assessed via an online and in-person questionnaire covering academic trajectory (grade point average, credits passed, curricular progression), sleep quality (PSQI), physical activity (IPAQ), tobacco use, Body Mass Index (BMI), and sociodemographic data. Bivariate associations were tested with chi-square, Pearson correlation, t-tests, and ANOVA, followed by generalized linear models for significant variables. Sports participation, but not IPAQ-measured physical activity, was significantly associated with academic progression, defined as advancing one grade level per year. Sleep indicators showed mixed, partly counterintuitive associations: poorer nocturnal sleep was linked to better progression, whereas better sleep quality predicted a higher percentage of credits completed. BMI was a modest positive predictor of credit completion, and underweight students performed significantly worse than overweight or obese peers. Tobacco use showed no significant associations. These findings suggest that sleep-related behaviors, sports participation, and, to a lesser extent, BMI, are relevant correlates of academic progress, supporting university health policies addressing sleep, physical activity, and nutrition.

Behavioral test batteries are increasingly used to characterize multiple functional domains in zebrafish, yet the potential impact of test sequence on behavioral outcomes remains poorly defined. Here, we systematically evaluated whether test order influences behavioral responses in a three-assay battery comprising the novel tank test (NTT), mirror-induced aggression (MIA), and social preference (SP) test. Adult zebrafish (Danio rerio) were exposed to all possible permutations of the three assays in a fully counterbalanced design, allowing assessment of order effects across locomotor, anxiety-like, aggression-related, and social behaviors. Test order produced modest and parameter-specific effects, primarily affecting locomotor activity in the NTT and social proximity in the SP assay. Time-course analysis revealed within-test behavioral dynamics, with limited evidence that test order modulates early adaptation or late engagement with the testing environment but does not alter overall temporal response profiles. Sex-dependent effects were assay-specific and most pronounced in the NTT, with no consistent sex differences observed in MIA or SP. To evaluate the global structure of behavioral variation, Principal Component Analysis (PCA) was performed across assays. Despite localized effects of test order, no clear multivariate separation between test sequences was observed, indicating that sequential testing does not produce distinct baseline phenotypes. Together, these findings support the robustness and reproducibility of multidomain behavioral batteries while highlighting the importance of standardized test-order reporting to improve cross-study comparability.

Operant paradigms are powerful tools to quantify motivation and reward. Traditionally, operant conditioning research has been limited to food and drug reinforcers. Recent advances in commercially available operant equipment, however, allow for the quantification of social motivation. These operant assays are an improvement over commonly used social preference tasks, as they enable direct measurement of the effort and motivation driving social behavior. Based on a design by Venniro et al. (2020), the MedPC social operant boxes modify the traditional operant box setup for social interactions. The experimental rat can lever-press to raise a door for an interaction with a target rat behind a porous barrier. These social operant boxes have been widely adapted to test social behavior in adult and adolescent rodents and investigate how a range of conditions (e.g. stress, drug taking, etc.) affect social motivation. However, there is a gap in assessing maternal motivation for pups during the postpartum period, despite ample evidence that postpartum social behavior is highly relevant for offspring health outcomes. Here, we detail 3D-printed modifications to the standard Med PC social operant boxes to adapt the social target chamber to safely house neonatal pups. We have also developed testing protocols to assess motivation during the limited postpartum period. These data demonstrate that, with simple modifications to social operant chambers and testing protocols, the field can implement advanced behavioral approaches to directly assess maternal motivation.

While hamsters are commonly housed in groups within pet shops in Japan, small cages are often thought to restrict their physical activity, leading to arguments that larger cages should be provided. Although previous research has investigated how cage size and running wheel availability influence activity levels in individually housed hamsters, no studies to date have examined these specific effects in a social housing context. Therefore, this study investigated how cage size and the presence of a running wheel affect the activity levels of individual hamsters using the group-housing conditions with five hamsters. Video recordings were captured for 24 hours across four distinct cage environments using a camera installed directly above each cage. From these recordings, the distances traveled on both the cage floor and the running wheel were calculated for each hamster and statistically analyzed. The results revealed that overall activity levels were significantly higher in cages equipped with a running wheel than in those without. Although cage size did not yield a statistically significant difference, a marginal trend toward higher activity in larger cages was observed.

Chorus howls play a central role in wolves' (Canis lupus) social communication, shaping both intra- and inter-pack interactions. While synchrony has been shown to influence perceived threat from outgroups in birds and humans, whether the synchrony of an outside pack's chorus howl plays a role in wolf pack behaviour remains unclear. This study investigated behavioural responses of 19 zoo-housed wolf packs to playbacks of synchronous and asynchronous chorus howls from unfamiliar packs, in comparison to a dove control call. We hypothesized that packs would show stronger intra-pack cohesion and greater spatial withdrawal in response to more synchronous, and thus cohesive - and potentially more formidable - sounding vocalisations. Alternatively, asynchronous howls might elicit stronger responses due to their ambiguity and the "Beau Geste" effect (where asynchronous signals make it harder to infer the number of callers). Synchronous howls resulted in more distance from the sound source and less activity compared with asynchronous howls. This could be interpreted as a more cautious response towards a perceived threat. However, since there was no effect on pack-directed behaviour and no difference between the synchronous and control sound, lower interest or motivation to engage with the stimulus cannot be excluded. In contrast, wolves spent more time near the sound-source after hearing asynchronous howls, approached it more, and oriented towards it for longer compared to synchronous playbacks, a pattern consistent with threat-investigation. In how far these reactions result from a more difficult localisation of synchronous howls or more ambiguity in pack size in the asynchronous playbacks, as well as how this translates to wild wolf behaviour, remains to be investigated.

Animal personality is the expression of consistent among-individual variation in a behavioural trait across time and context. The application of this theory to behavioural research provides a valuable framework to investigate the among- and within-individual variation in behaviour. The animal personality theory is particularly relevant to understand and diagnostic fish welfare. It can be integrated within the nature-based welfare framework, which emphasizes the expression of species-specific behaviours and the maintenance of consistent behavioural patterns over time. Because behavioural trait such as boldness may be closely linked to other functional phenotypes, such associations reflect the broader concept of animal personality, whereby a behavioural trait can covary with another phenotypic trait to support an adaptive responses to environmental conditions. Although these relationships are both species and context dependent, they are consistently shaped by environmental conditions and environmental complexity tend to promote species specific behaviours and reduce maladaptive traits. Here we examined how structural environmental complexity shapes personality in Arctic charr and their covarying functional phenotypes, specifically growth rate and brain size. We propose that environmental complexity promotes alternative behavioural and functional phenotypes through multivariate phenotypic plasticity. We found that environmental complexity did not influence mean boldness between both treatments, but repeatability of boldness in the complex environment was remarkably consistent over a longer-term period, while estimates collapse after seven days in the plain treatment. Our findings are a clear evidence that environmental complexity foster stable behavioural trait expression and that a plain environment may suppress personality. Our results provide compelling evidence that behavioural structure and dynamics are embedded within patterns of behavioural variance. Although we found no support for behavioural covariation or associations with growth rate and brain size, we suggest that the animal personality framework may offer a valuable approach for diagnosing fish welfare issues through the partitioning of behavioural variance.

Intergroup conflict is one of the most significant issues in human society. In the 1950s, Sherif et al. reported that intergroup conflict could be artificially induced in boys through intergroup competition with tug-of-war and ball games. Since this iconic study, researchers have developed various experimental methods to replicate intergroup competition and/or conflicts. However, although intergroup conflicts in wild animals are often reported, it has been difficult to establish a situation of intergroup conflict in laboratory rodents that is discriminable from aggressive behavior individually. In this study, we established a novel experimental paradigm for intergroup competition in mice in which the members of each group shared objectives and tasks. Adult male ICR/Jcl mice were housed in groups of six, divided into two teams of three and repeatedly performed a competitive Tsunahiki task (tsunahiki means tug-of-war in Japanese). The competitive Tsunahiki task was conducted in an open field divided into two experimental fields, with three ropes stuck to a wall separating the fields. The mice were required to pull two ropes out faster than their opponent team to win, and only the winners could proceed to the reward area separated by a guillotine door. We demonstrated that the experience of the competitive Tsunahiki task induced attack bites selectively toward members of the other team (out-group members). Our findings suggest that intergroup competition induces intergroup conflict in mice, providing a technical breakthrough in elucidating the detailed neuroscientific mechanisms underlying intergroup conflict.

Abstract Introduction: Despite advances in pain management, effective analgesics in pain situations remain elusive. Opioids and non-opioids carry risks of neurotoxic and psychedelic effects with adverse physiological outcomes. Indian instrumental music (IIM) mitigates subacute pain by rewiring neurochemical synergy as an evidence-based, non-invasive, non-pharmacological system to mitigate pain. Objective: Investigating therapeutic efficacy of IIM in mitigating subacute pain by analyzing behavioral, peripheral, and central neurochemical re-tuning. Methods: Mice were divided into Control, Pain, Pain+Music, and Music groups. Pre-treatment behavioral parameters were compared with those observed after 14 days IIM exposure. Evaluations included nociceptive latencies (hot-plate/tail-flick), locomotion (Open Field Test), and anxiety (Elevated Plus Maze). Molecular analyses quantified peripheral neuropeptides (SP, NK-1R, CGRP), serum cortisol, spinal neurotrophic factor, neurotransmitters (glutamate, GABA, dopamine (DA), 5-HT), BDNF, and mRNA expression of BDNF, Ntrk1R/2R, and D1R in cortex, thalamus, hippocampus and hypothalamus. All procedures adhered to IAEC guidelines. Results: IIM yielded 3.9-4.4-fold antinociceptive improvements, 3.3-fold locomotor restoration, and 3.6-4.9-fold anxiolysis. 14 days IIM exposure reduced peripheral nociceptive-neuropeptides 1.3-2.0-fold (SP, NK-1R, CGRP), serum cortisol 1.3-fold, and spinal glutamate, serotonin levels 1.5- and 1.3-fold. An enhanced expression of spinal GABA, DA about 1.5-fold, and BDNF by 1.3-fold was observed after music listening. Brain-region-specific differential mRNA-expression at cortex, thalamus, hypothalamus and hippocampus revealed the neuromodulatory impact of rhythmic music in a formalin-induced murine pain-model. Conclusion: Gross reduction of pain parameters demonstrates therapeutic potential of IIM as multilevel neuromodulator to suppress the multidimensional stressor, pain, via peripheral desensitization, spinal E-I balance, and differential calibration of BDNF/Trk/D1R plasticity at specific brain-regions. Keywords: Pain, Non-Pharmacological Method, Indian Instrumental Music (IIM), Behavior, Neurotransmitters, Neuroplasticity, mRNA Expression.

Long-term opioid therapy is limited by analgesic tolerance and opioid-induced hyperalgesia, but the roles of genetic background, sex, and drug exposure remain unclear. We used 20 inbred strains from the Hybrid Rat Diversity Panel to examine thermal sensitivity, oxycodone analgesia, tolerance, and hyperalgesia-like changes following voluntary intravenous oxycodone or saline self-administration. Rats underwent tail-immersion testing before self-administration (Pre-SA) and after self-administration (Post-SA). Oxycodone analgesia was assessed using the percent maximum possible effect time course and the corresponding area under the curve. Pre-SA thermal sensitivity differed across strains and between sexes, and Pre-SA oxycodone analgesia also differed across strains. Oxycodone self-administration produced a sex-dependent increase in thermal sensitivity that was most evident in males. During Post-SA testing, oxycodone self-administering rats showed reduced analgesic responsiveness compared with saline controls, and the magnitude of this difference varied across strains. Within-strain Pre-SA-to-Post-SA comparisons identified tolerance-like reductions in several strains. Across strains and sexes, oxycodone self-administering rats showed a greater Pre-SA-to-Post-SA reduction in analgesic responsiveness than saline controls, consistent with analgesic tolerance. Total oxycodone intake was not associated with tolerance at either the strain-mean or individual-animal level. Heritability estimates were higher for thermal sensitivity and analgesia (H2 {approx} 0.28-0.40) than for changes in thermal sensitivity and tolerance (H2 {approx} 0.18-0.27). These findings demonstrate strain variation in thermal sensitivity and oxycodone analgesia, sex-dependent hyperalgesia-like effects, and reduced analgesic responsiveness following voluntary oxycodone intake.

Honey bees can swim on the water surface toward dark regions, a behavior known as scototaxis that may facilitate escape from water. Although this behavior has been reported in both honey bees and solitary bees, variation among honey bee species remains poorly understood. We compared scototaxis during swimming in four honey bee species representing two nesting types: open-nesting (Apis florea and A. dorsata) and cavity-nesting (A. cerana and A. mellifera). Individual bees were released into a water-filled arena containing a dark sector, and their landing angles were recorded. All species exhibited significant orientation toward the dark sector. However, open-nesting species showed significantly stronger orientation than cavity-nesting species. No significant differences were detected between replicate colonies within species or between species within the same nesting type, whereas differences between nesting types were highly significant. Hierarchical clustering based on orientation strength placed Osmia, a solitary cavity-nesting bee from our previous study, in the same behavioral cluster as the two open-nesting Apis species rather than the cavity-nesting honey bees. We also measured swimming duration, distance, and velocity, but found no consistent differences between nesting types. These results demonstrate substantial interspecific variation in swimming scototaxis. The behavioral clustering is consistent with the hypothesis that strong scototaxis represents an ancestral trait that has been reduced in the derived A. cerana/A. mellifera lineage.

Traditionally, complex behavioral tasks in mice have relied upon water restriction as an external motivator to increase task engagement. However, citric acid (CA) water, the technique whereby water is given ad libitum but made sour by the addition of CA, has emerged as an alternative to typical water restriction. Evidence suggests CA water can effectively motivate task performance while improving animal welfare in alignment with the 3Rs of animal research and reducing experimenter labor. While promising, the applicability of CA water in mice remains incompletely characterized with higher concentrations only tested in rats and "ramping" schedules, where mice progress to increasingly higher concentrations, indirectly examined. Here, we evaluate four CA concentration/schedule combinations for their effects on mouse health (weight changes, home cage behaviors, fecal counts) and motivation to drink regular water (lick counts, drinking behaviors) in female and male C57BL/6J mice. We find that a schedule ramping from 1% to 2% CA after one week is the easiest for mice to adapt to and sustained 2% CA use maintains robust lick counts for at least 5 weeks. Additionally, CA has been directly characterized for wheel-turning and touchscreen tasks, but not virtual reality (VR) tasks, an increasingly popular class of behavioral experiments. Therefore, we also assess how 2% CA affects motivation and task performance in two VR treadmill tasks (running and stopping task). We find that CA water does not improve task performance above that of mice given regular water, but does limit competing motivations and produce more uniform, reward-motivated behavior.

RationalePsychedelic compounds such as psilocybin have attracted growing interest for their potential therapeutic effects in psychiatric disorders, with improvements in cognitive flexibility proposed as a possible mechanism of action. However, the effects of psychedelics on cognitive flexibility remain poorly understood. ObjectiveThis study aimed to examine the acute and post-acute effects of psilocybin (0.1, 0.3, 1 mg/kg) and lysergic acid diethylamide (LSD, (0.02, 0.04, 0.08 mg/kg) on cognitive flexibility in male rats. MethodsThis was tested using two variants of the probabilistic reversal learning task (PRLT): a touchscreen-based operant task and a more ethological foraging-based task. ResultsIn the touchscreen PRLT, acute psilocybin disrupted task engagement, with animals completing fewer trials and showing increased trial initiation latency, although psilocybin also showed a trend toward faster initial rule acquisition. However, psilocybin did not significantly alter the number of rule changes achieved, a canonical measure of cognitive flexibility, or feedback sensitivity. LSD similarly produced limited acute effects, although the highest dose reduced lose-shift probability, suggesting decreased sensitivity to negative feedback under some conditions. Post-acute effects of psilocybin were minimal in both PRLT variants and, where LSD effects were observed these occurred across different doses and timepoints without a consistent pattern. ConclusionsOverall, these findings suggest that serotonergic psychedelics do not robustly enhance reversal learning in these paradigms and that apparent learning effects may reflect transient disruptions in task engagement rather than improvements in cognitive flexibility. These results also highlight potential limitations of these PRLT paradigms for detecting psychedelic-induced changes in cognitive flexibility in rodents.

IntroductionExecutive functions (EF) are higher-order cognitive processes essential for academic performance in university settings. Although there is extensive research on EF training in children, studies in young adults are scarce, particularly those involving interventions tailored to specific needs. ObjectiveTo evaluate the effect of the University Executive Network-Training Program (NExT-U), based on specific needs, on the executive functioning of Cuban university students. MethodologyA quasi-experimental study with a non-equivalent control group and pretest-posttest measurements. Participants were 27 second-year Psychology students (74% female; mean age = 19 years). The experimental group (n=7) received three training sessions focused on Conscious Regulation of Behavior, Decision-Making, Emotional Regulation, and Monitoring of Responsibilities, identified through an initial assessment using the UEF-1 Scale. The control group (n=20) continued with their usual academic activities. Non-parametric analyses and the residual gain method were employed. ResultsThe experimental group showed significant improvements in Conscious Regulation of Behavior (p = .026; r = .51), Emotional Regulation (p = .030; r = .49), and the Supervisory Attention System (p = .046; r = .44), with large effect sizes. The control group experienced no significant changes in any of the functions evaluated. ConclusionsA brief, personalized program can enhance specific executive functions in university students, demonstrating cognitive plasticity in young adults. The findings support the design of contextually relevant interventions to strengthen transversal competencies in higher education.

Planaria exhibit remarkable regenerative ability, including the capacity to regrow complete heads and brains after decapitation. Here, we re-investigated whether regenerated planaria can preserve learned avoidance behavior, a phenomenon that has been reported previously but has been difficult to study due to unreliable experimental protocols. Using a light-to-food associative conditioning paradigm, planaria were trained to override their normal photophobic preference and then decapitated. Following a two-week regeneration period, behavioral responses to the conditioned stimulus were re-evaluated. Results indicated that the majority of regenerated planaria retained the learned response, supporting a model in which behavioral patterns can regenerate as well as anatomical patterns. By establishing a consistent, low-cost, and effective protocol for studying memory persistence through regeneration, such work may help inform future research on memory loss, resilience, and recovery in neurodegenerative diseases.

Cruising speed is a key factor affecting prey-search efficiency and migration range in continuously swimming animals. Tunas and lamnid sharks (e.g., white sharks) have convergently evolved traits for high-speed cruising, including the ability to maintain slow-twitch, aerobic red muscle (RM) warmer than ambient water, known as RM endothermy. Despite their well-known high cruising speeds, kinematic features underlying their elevated speeds remain unclear. Swim speed is the product of tailbeat frequency (TBF; Hz) and stride length (SL, the absolute distance traveled per tailbeat; m). RM endothermy is expected to elevate TBF by enhancing muscle contraction performance. Furthermore, within RM-endothermic fishes, tunas and lamnid sharks may exhibit distinct kinematic features because of differences in caudal fin morphology and tailbeat amplitude. Here, we compiled kinematic parameters from 20 fish species, including five RM-endothermic species, measured in the wild using animal-borne sensors. Comparative analyses showed that, for a given body mass and water temperature, RM-endothermic fishes exhibited 1.9 times higher cruising speed and TBF than ectothermic fishes, while SL remained similar. Within RM-endothermic fishes, tunas exhibited 2.3 times higher TBF than similar-sized lamnid sharks, whereas lamnid sharks showed 1.7 times longer SL than similar-sized tunas. These results indicate that RM endothermy is generally associated with higher TBF, while significant kinematic differences remain between tunas and lamnid sharks. This divergence may be partly explained by the greater caudal fin area and tailbeat amplitude in lamnid sharks. It may also reflect contrasting skeletal types of teleosts and elasmobranchs, which potentially influence body stiffness and swimming kinematics.

Environmental predictability influences the value of information acquired through experience, yet relatively little is known about how instability in resource characteristics influences behavioural organisation during foraging. We tested whether repeated changes in floral orientation, a manipulation of environmental predictability, affect pollen foraging in bumblebees (Bombus terrestris) by exposing naive workers to either stable floral conditions (single flower orientation) or repeated inter-trial changes in flower orientation (three alternating flower orientations), under constant resource availability. We quantified pollen collection, foraging efficiency, revisitation behaviour, floral coverage and sonication behaviour across three successive foraging trials of either constant or variable flower orientation, before assessing performance in a common post-treatment preference test in which bees were offered all three flower orientations and higher pollen rewards. Environmental instability altered the organisation of foraging behaviour. Bees exposed to unstable floral conditions progressively reduced flower revisitation behaviour and were less likely to perform sonication, although floral coverage, defined as the number of unique flowers visited, remained unchanged. Contrary to our predictions, instability had only weak immediate effects on pollen acquisition and foraging efficiency compared to bees tested under stable conditions. However, previous exposure to instability generated carry-over effects in the common post-treatment preference test. Bees previously exposed to unstable conditions were significantly less likely to return with pollen and consequently collected less pollen overall than bees previously exposed to stable conditions. Our results demonstrate that environmental instability can influence pollen foraging in ways that are not captured by immediate measures of performance. Although behavioural adjustments appeared to buffer short-term consequences during repeated foraging trials, carry-over effects were evident when bees were later tested in a common high-reward, multi-orientation floral array. These findings highlight the importance of considering both behavioural flexibility and carry-over effects when evaluating how organisms respond to changing environments.

Understanding the neurobiological systems that regulate alcohol cue-induced craving is of utmost importance for the development of novel intervention strategies for alcohol use disorders (AUDs). However, although a human experimenter is required to conduct alcohol self-administration studies in the lab, the cues associated with the experimenter are seldom if ever factored into the experimental design. Thus, although we have learned much to date about alcohol cue-induced behavior and neurobiology, and in particular about discrete cues presented many times throughout a single daily alcohol self-administration session, we know relatively little about how responses to alcohol availability cues might predict subsequent alcohol consumption. For the current experiment, mice were exposed daily to auditory cues that preceded 2 hours of alcohol or water access using drinking-in-the-dark (DID) methods. An additional control group experienced cues but were not otherwise manipulated. Importantly, cues were initiated remotely from outside the animal facility, avoiding the experimenter being the first cue predicting ethanol availability. Head direction, location in the home cage, and movement velocity were the primary variables on interest. Surprisingly, during the cue period, there were no significant differences between groups in any of these measures, despite meaningful alterations over days. However, we observed many significant correlations between behaviors and drinking variables. First, we observed significant positive associations between ambulatory velocity during cues and subsequent total alcohol (R2=0.14; p<0.0001) and total water (R2=0.12; p=0.0002) consumption, but only in females. We also observed a significant positive relationship (R2=0.25; p<0.0001) between the amount of time oriented toward the sipper port during the auditory cues and the average rate of subsequent alcohol consumption (i.e. front-loading), but only in females. In males, head direction was found to be positively associated with subsequent total water consumption (R2=-0.21; p<0.0001), but not alcohol (R2=-0.01; p=0.2267). We also observed a significant negative relationship (R2=-0.15; p<0.0001) between proximity to the sipper during the cue period and subsequent total 2-hour alcohol intake in males. Although these associations were modest in strength, they suggest potential sex-specific behavioral predictors of alcohol consumption that are regulated by different neural dynamics.